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Expression and characterization of a novel isocitrate dehydrogenase from Streptomyces diastaticus No. 7 strain M1033

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Abstract

Isocitrate dehydrogenase (IDH) is one of the key enzymes in tricarboxylic acid cycle, widely distributed in Archaea, Bacteria and Eukarya. Here, we report for the first time the cloning, expression and characterization of a monomeric NADP+-dependent IDH from Streptomyces diastaticus No. 7 strain M1033 (SdIDH). Molecular mass of SdIDH was about 80 kDa and showed high amino acid sequence identity with known monomeric IDHs. Maximal activity of SdIDH was observed at pH 8.0 (Mn2+) and 9.0 (Mg2+), and the optimal temperature was 40 °C (Mn2+) and 37 °C (Mg2+). Heat-inactivation studies showed that SdIDH remained about 50 % activity after 20 min of incubation at 47 °C. SdIDH displayed a 19,000 and 32,000-fold (k cat/K m) preference for NADP+ over NAD+ with Mn2+ and Mg2+, respectively. Our work implicate that SdIDH is a divalent metal ion-dependent monomeric IDH with remarkably high coenzyme preference for NADP+. This work may provide fundamental information for further investigation on the catalytic mechanism of monomeric IDH and give a clue to disclose the real cause of IDH monomerization.

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Acknowledgments

This research was supported by funds from the National Natural Science Foundation of China (31170005; 30870062), Specialized Research Fund for the Doctoral Program of Higher Education of China (20113424110004), the Fund of State Key Laboratory of Genetics Resources and Evolution from Kunming Institute of Zoology (Chinese Academy of Sciences, CAS) (GREKF11-07), the National High Technology Research and Development Program (“863” Program: 2012AA02A708), Anhui Provincial Key Laboratory of the Conservation and Exploitation of Biological Resources and Program for Innovative Research Team in Anhui Normal University.

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Authors and Affiliations

  1. Key Laboratory of Molecular Evolution and Biodiversity and Institute of Molecular Biology and Biotechnology, College of Life Sciences, Anhui Normal University, No.1 Beijing East Road, Wuhu, 241000, Anhui, People’s Republic of China

    Bei-Bei Zhang, Peng Wang, Ao Wang, Wen-Cai Wang, Wang-Gang Tang & Guo-Ping Zhu

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  1. Bei-Bei Zhang
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  2. Peng Wang
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  3. Ao Wang
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  4. Wen-Cai Wang
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  6. Guo-Ping Zhu
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Correspondence to Guo-Ping Zhu.

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B.-B. Zhang and P. Wang contributed equally to this work.

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Zhang, BB., Wang, P., Wang, A. et al. Expression and characterization of a novel isocitrate dehydrogenase from Streptomyces diastaticus No. 7 strain M1033. Mol Biol Rep 40, 1615–1623 (2013). https://doi.org/10.1007/s11033-012-2210-y

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  • DOI: https://doi.org/10.1007/s11033-012-2210-y

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